Abstract
Objective: Although acute and chronic roles of microvascular smooth muscle cells as effectors of pulmonary vascular resistance and remodeling are well appreciated, relatively little is known regarding the direct effects of neurohumoral agents on these cells. We recently reported that microvascular smooth muscle cells isolated from distal rat lung (RPC) express mRNA and binding sites for 5HT2 receptors. The objective of the current study was to determine if protein kinase C (PKC) affected 5HT-induced changes in intracellular calcium and phosphoinositide metabolism in RPC.
Methods: 5HT-induced changes in intracellular calcium ([Ca2+]i) in single RPC were determined microspectrofluorometrically using the calcium-sensitive dye, Fura-2.
Results: Phorbol 12-myristate 13-acetate (PMA: 100 nM) caused a rapid desensitization of 5HT-induced increases in [Ca2+]i. Staurosporine, a putative PKC inhibitor, abolished the PMA-induced desensitization. Downregulation of PKC with prolonged (24 hr) PMA exposure also abolished subsequent PMA-induced desensitization of 5HT response. Neither short- nor long-term exposure of RPC to PMA affected binding of [125I]LSD. Activation of PKC by PMA was associated, however, with complete inhibition of 5HT-induced increases in intracellular inositol monophosphate.
Conclusions: These data are consistent with PKC causing desensitization of 5HT2 receptors by affecting elements of signal transduction and uncoupling receptor-G protein complex from phospholipase C in RPC.